This project is designed to provide information about the organization of neuronal systems in the mammalian spinal cord that are involved in the neural control of movement. There are three current sub-projects: 1) studies of synaptic transmission; 2) studies of the neural circuits that produce coordinated locomotion in a primitive amphibian; and 3) studies of the quantitative morphology of motoneurons in the neonatal mouse spinal cord. All of these studies use in vitro preparations of brain stem slices or isolated en bloc spinal cord of neonatal mice or amphibians. Experiments on low-frequency synaptic depression of group Ia monosynaptic excitatory postsynaptic potentials (EPSPs) produced in lumbosacral spinal motoneurons by repetitive stimulation of dorsal roots were completed in FY2002. During FY 2003, we completed data analysis and published a full research report of post-natal age-related changes in short-term depression between postnatal ages of two (P2) to twelve (P12) days. Using an empirical synaptic release model described in previous Annual Reports, our data suggest that transmitter release fraction systematically decreases with postnatal age, while the coefficients and decay time constants of facilitation and augmentation processes change in more complex ways. A research report on age-related changes in the effects of altered external calcium concentration and of blockade of specific types of calcium channels is now being completed for publication in the Journal of Neurophysiology. We also completed tests of our synaptic release model in collaboration with scientists at the Australian National University in Canberra, using data from the giant end bulb of Held in the brain stem auditory nucleus, AVCN. The long range goal of a second project is to elucidate spinal cord mechanisms that control rhythmic walking movements in a relatively primitive amphibian, Necturus maculosus. In order to design appropriate electrophysiological experiments to examine the spinal circuits involved in locomotor movements in Necturus, we required detailed information about the location and nature of its spinal neurons and synaptic interconnections. We have now completed an extensive study of the anatomy of the cervical spinal cord in Necturus, which had never before been described. Motoneurons, interneurons, and primary afferents were labeled with fluorescent tracers and studied by confocal microscopy. Cell bodies of both motoneurons and interneurons lie in a gray neuropil along the lateral and ventral borders of the gray matters, immediately adjacent to the white matter into which their dendrites project. Remarkably, the central region of gray matter is virtually empty except for glial cell bodies. Labeling lateral white matter tracts fill interneurons located on both sides of the cord. All crossing axons and dendrites travel in a thin sheet beneath the central canal region. At the lateral border of the Necturus cord, distal terminations of motoneuron dendrites expand into elaborate, parasol-like tangles of thin branches in a sub-pial plexus just beneath a thin layer of glia. Electron microscopy showed that the glia limitans layer contains arrays of tubular structures that have not to our knowledge been described before. A research report of this work will be submitted in early FY2004, when electrophysiological experiments will begin. Analysis of data from 20 fully-reconstructed motoneurons in two age groups of neonatal mice have been largely completed in collaboration between this laboratory and scientists at the Krasnow Institute of George Mason University. The results from intracellular labeling of individual motoneurons indicate that somatic size increase only modestly between 2 and 11 days postnatal (P2 - P11), while there is a large increase in the length and surface area of entire trees between these ages, with little change in their topological structure. A corollary study of the sizes and packing density of neonatal motoneurons labeled by retrograde transport of fluorescent markers from ventral roots to large numbers of motoneurons indicate that there is indeed a significant change in average somatic sizes between P2 and P11, as well as a decrease in packing density. These quantitative data show that intracellular techniques are biased toward larger neurons, which was anticipated, but provide for the first time a quantitative estimate of the magnitude of this bias. The data will form part of an extensive data base being developed as part of the Human Brain Project that is designed to permit scientists anywhere to access and analyze morphological data on many types of neurons.